DESCRIPTION: The main goal of this research proposal is to identify the factor(s) that trigger initial induction of craniofacial cartilage by neural crest. The first aim is to set up in vitro assay cultures in which premigratory neural crest cells are first proven to be the only occupants of the cultures and then tested for lack of ability to form cartilage prior to addition of the factor(s) of interest. The first aim will utilize better antibody markers than previous investigations for a more rigorous validation of the test system. Secondly, actions of BMP proteins will be tested in permissive conditions and when antagonized by noggin or other molecules. In addition, the required induction by ectodermal cells will be tested in co-culture when inductive expression of BMP is blocked. Third, other genes such as endothelin-1, TGF-beta, PDGF or Sfrp-2 (for which a role in cartilage induction has been suggested by other work) will be tested for positive actions, and using dominant-negative receptors that are added to neural crest by electroporation for specific inhibition of specific factors. Fourth, trunk crest capabilities for chondrogenesis will be tested by grafting to various cranial sites. When trunk and cranial tissues differ, they will be further analysed by looking for presence or absence of receptors for the key inductive factors. There are no special training activites planned besides those related to the candidate's research and the regular laboratory meetings, journal clubs, etc. of Dr. Bronner-Fraser's group (none would be expected for a young scientist at Dr. Knecht's level of experience). The candidate expects regular interactions with other Cal Tech laboratories, especially those of Drs. Scott Fraser and David Anderson, and she will make use of the Cal Tech imaging center.